PDBsum entry 1kea

Hydrolase

PDB id: 1kea

Contents

Protein chain

217 a.a. *

Ligands

ACT ×4

SF4

Metals

_ZN

_CL ×2

Waters ×200

* Residue conservation analysis

PDB id:

1kea

Name:

Hydrolase

Title:

Structure of a thermostable thymine-DNA glycosylase

Structure:

Possible g-t mismatches repair enzyme. Chain: a. Synonym: mismatch glycosylase. Engineered: yes

Source:

Methanothermobacter thermautotrophicus. Organism_taxid: 145262. Expressed in: escherichia coli. Expression_system_taxid: 562

Resolution:

2.00Å R-factor: 0.211 R-free: 0.248

Authors:

C.D.Mol,A.S.Arvai,T.J.Begley,R.P.Cunningham,J.A.Tainer

Key ref:

C.D.Mol et al. (2002). Structure and activity of a thermostable thymine-DNA glycosylase: evidence for base twisting to remove mismatched normal DNA bases. J Mol Biol, 315, 373-384. PubMed id: 11786018 DOI: 10.1006/jmbi.2001.5264

Date:

14-Nov-01 Release date: 23-Jan-02

Protein chain

P29588  (MIG_METTF) -  Thymine/uracil-DNA glycosylase from Methanothermobacter thermautotrophicus

Seq: 221 a.a.

Struc: 217 a.a.

Enzyme reactions

• Enzyme class 2: E.C.3.2.2.- - ?????
• Enzyme class 3: E.C.3.2.2.29 - thymine-DNA glycosylase.

Note, where more than one E.C. class is given (as above), each may correspond to a different protein domain or, in the case of polyprotein precursors, to a different mature protein.

DOI no: 10.1006/jmbi.2001.5264

J Mol Biol 315:373-384 (2002)

PubMed id: 11786018

Structure and activity of a thermostable thymine-DNA glycosylase: evidence for base twisting to remove mismatched normal DNA bases.

C.D.Mol, A.S.Arvai, T.J.Begley, R.P.Cunningham, J.A.Tainer.

ABSTRACT

The repair of T:G mismatches in DNA is key for maintaining bacterial restriction/modification systems and gene silencing in higher eukaryotes. T:G mismatch repair can be initiated by a specific mismatch glycosylase (MIG) that is homologous to the helix-hairpin-helix (HhH) DNA repair enzymes. Here, we present a 2.0 A resolution crystal structure and complementary mutagenesis results for this thermophilic HhH MIG enzyme. The results suggest that MIG distorts the target thymine nucleotide by twisting the thymine base approximately 90 degrees away from its normal anti position within DNA. We propose that functionally significant differences exist in DNA repair enzyme extrahelical nucleotide binding and catalysis that are characteristic of whether the target base is damaged or is a normal base within a mispair. These results explain why pure HhH DNA glycosylases and combined glycosylase/AP lyases cannot be interconverted by simply altering their functional group chemistry, and how broad-specificity DNA glycosylase enzymes may weaken the glycosylic linkage to allow a variety of damaged DNA bases to be excised.

Selected figure(s)

Figure 2.
Figure 2. Stereo views of the MIG structure, enzyme homology and DNA-interaction surface. (a) The MIG structure illustrating the two-domain architecture with the a-helices (purple) of the four-helix domain (top), and six-helix barrel domain (bottom) numbered sequentially. The a-helices and b-hairpin loop of the HhH motif (orange), the Fe[4]S[4] iron-sulfur cluster (large spheres), and the positions of key residues lining the active site at the domain interface are shown. (b) The MIG structural homology with other HhH glycosylases and combined glycosylase/AP lyases. The known structures of HhH DNA repair enzymes are superimposed on the structure of M. thermoformicicum MIG according to conserved structural elements within their six a-helix barrel domains. (c) The MIG molecular surface in the same orientation as shown in (a), and colored by electrostatic charge (color bar: red, -2.0 kT/e to blue, +2.0 kT/e). The positively-charged DNA-binding face of MIG is a conserved feature of the homologous HhH glycosylases and combined glycosylase/AP lyases and assists in orienting the DNA for nucleotide-flipping of target base lesions into the active site cleft. DNA (red tubes) is shown superimposed on the MIG surface from the homologous AlkA:DNA complex structure.

Figure 5.
Figure 5. Structurally implied active site chemistry for pure HhH glycosylase MIG. MIG interactions of Glu42 and Tyr126 with the O4, N3 and O2 positions of thymine facilitate glycosylic bond dissociation, while the vert, similar 90° clockwise twist and distortion enforced by the MIG thymine-binding pocket allows the three normally orthogonal O4', N-C1', and p electron orbitals to overlap. This orbital overlap facilitates catalysis by promoting the electron transpositions needed for glycosylic bond cleavage[16].

The above figures are reprinted by permission from Elsevier: J Mol Biol (2002, 315, 373-384) copyright 2002.

Figures were selected by an automated process.